The kinetics of rotavirus infection were examined at the ultrastructural and molecular level. As early as 6 hr postinfection, with a cytolytic bovine rotavirus (C486), particles could be seen in the cytoplasm. At times later than 6 hr postinfection (p.i.) the number of virus particles rapidly increased such that at 12–18 hr p.i. large numbers of particles were present in rough endoplasmic cysternae (RERc) and some particles were budding through the RERc. Concomitant with virus replication, as determined by electron microscopy and labeling of viral products, there was generalized destruction of all cytoplasmic organelles as well as the nucleus and inhibition of host cell macromolecular synthesis as measured by decreased incorporation of 3H-thymidine, 3H-uridine, and 3H-amino acids. In addition to inhibition of host cell DNA synthesis, there was also breakage of host cell DNA. The rapidity with which the virus shuts off host cell function and causes cell death is discussed in relationship to the speed with which rotavirus-induced diarrhea occurs in animals following infection with rotavirus.